Protection Of Protein Tyrosine Phosphatase 1B By Persulfides Against 9,10-Phenanthrenequinone-produced Reactive Oxygen Species In Vitro

抄録

　Redox signaling [e.g., PTP1B/EGFR signaling] through oxidation of cysteines in proteins is involved in maintaining intracellular homeostasis. Once SH group on PTP1B active cysteine undergoes oxidation by reactive oxygen species (ROS) to form reversible SOH, and further irreversible SO₃H, PTP1B activity is inhibited, resulting in EGFR phosphorylation. We found that 9,10-phenanthrenequinone (9,10-PQ), a major quinone in PM_2.5, produced ROS excess through its redox cycling. While 9,10-PQ-produced ROS drastically decreased recombinant PTP1B activity by irreversible oxidation of Cys, exposure of A431 cells to 9,10-PQ did not markedly inhibit intracellular PTP activity and caused transient EGFR activation, suggesting that the cells have a convenient system to block further oxidation of PTP1B. Hence, we investigated the involvement of persulfide (SSH) with high-nucleophilicity in the ROS-dependent modification of PTP1B caused by 9,10-PQ.

　To generate S-sulfhydrated Cys, PTP1B was reacted with sodium disulfide (Na₂S₂). As a result, Cys215, PTP1B active site, was converted to SSH, which was reversible. Although excessive ROS cause oxidation of SH to irreversible SO₃H, we postulated that reaction products of SSH with ROS are SSOH, SSO₂H, and SSO₃H that could be reduced by dithiothreitol (DTT) regenerating SH. Consistently, we succeeded in detecting SSOH conversion at Cys215 during a reaction of PTP1B with H₂O₂ or 9,10-PQ in Na₂S₂ presence. As expected, SSOH was reduced to SH by DTT and PTP1B activity was restored. Under these conditions, oxidized PTP1B was decreased, suggesting a reversibility of SSO_nH (n = 1~3) associated with PTP1B activity recovery.

　This study indicates, for the first time, that persulfide-mediated S-sulfhydration at Cys215 of PTP1B appears to protect the active site against oxidation by excessive ROS. In other words, persulfides might repress further irreversible oxidation of protein thiols.